Malaria remains a major global infectious disease, largely affecting people living in resource poor environments, and is one of the most important causes of childhood mortality. Drug-resistance is constantly undermining the usefulness of antimalarial regimens. There is an urgent need for the development of new therapeutic strategies. Plasmodium falciparum parasites utilize multiple ligand-receptor interactions for the invasion of human erythrocytes. Much work has focused on the characterization of parasite ligands, with the goal of developing them as vaccine candidates. Largely due to the genetic intractabiliy of enucleated human erythrocytes, the function of host human erythrocyte receptors in ligand-receptor interactions has not been comprehensively assessed. We have developed an approach combining an in vitro erythrocyte culture system, which supports P. falciparum invasion and growth, with lentiviral transduction to generate genetically modified erythrocytes. The best characterized ligand-receptor interaction in P. falciparum is that of the ligand EBA-175 and its receptor glycophorin A, with EBA-175 being developed as a vaccine candidate. Using this system, we have achieved knockdown in expression of glycophorin A, genetically demonstrating that it is required for efficient strain-specific parasite invasion. We hypothesize that targeting a limited number of erythrocyte receptors will be sufficient to abrogate P. falciparum invasion in all P. falciparum parasite lines. This project will functionally analyze erythrocyte determinants of the invasion process of P. falciparum. We will use 1) a loss-of- function approach to establish a hierarchy amongst putative erythrocyte receptors for invasion, 2) a combinatorial knockdown approach to identify minimal sets of erythrocyte surface determinants that are essential for invasion and 3) a genetic screen to identify human blood group antigens that are novel determinants of invasion. In the long-term we hope that our studies will provide a functional understanding of critical ligand-receptor interactions for P. falciparum invasion of erythrocytes to inform vaccine development and the design of host-targeted therapeutics.